Errors during cell division can lead to genomic instability and aneuploidy, contributing to the generation of cancer and birth defects. A select group of kinases has been found to orchestrate mitosis. In particular, members of the cyclin-dependent kinase, Aurora, Polo, and NIMA/Nek families phosphorylate substrates in chromatin and at the spindle apparatus to regulate events during cell division (Nigg, Nat. Rev. Mol. Cell. Biol. 2: 21-32 (2001)).
Not surprisingly, histones are major targets of mitotic kinases. For example, histone H3 is extensively phosphorylated at serine-10 during mitosis and meiosis (Hendzel, et al., Chromosoma 106:348-360 (1997); Prigent, et al., J. Cell Sci. 116:3677-3685 (2003)). The function of this modification is debated, but it may facilitate chromatin condensation or the release of cohesin and ISWI chromatin-remodeling ATPases (Van Hooser, et al., J. Cell Sci. 111:3497-506 (1998); Andrews, et al., Curr. Opin. Cell Biol. 15:672-683 (2003); Prigent, et al., J. Cell Sci. 116:3677-3685 (2003); Swedlow, et al., Mol. Cell. 11:557-569 (2003)). A Tetrahymena strain with histone H3 mutated at serine-10 showed perturbed chromatin condensation and abnormal chromosome segregation during meiosis and mitosis (Wei, et al., Cell 97:99-109 (1999)), while a similar mutation in S. cerevisiae had no such effect (Hsu, et al., Cell 102:279-291 (2000)). Therefore, histone phosphorylation at serine-10 has an important role in mitosis but the extent to which it is required appears species-dependent, perhaps because of redundancy provided by other mitotic histone modifications ((Hsu, et al., Cell 102:279-291 (2000)). In fact, a number of highly conserved serine and threonine residues that might be phosphorylated are found in the core histones of all eukaryotes. Mitotic phosphorylation of threonine-11 (Preuss, et al., Nucleic Acids Res. 31:878-885 (2003)) and serine-28 (Goto, et al., J. Biol. Chem. 274:25543-25549 (1999)) of H3 has been reported.
The identities of protein kinases that phosphorylate the histones during mitosis in vivo remain somewhat uncertain. The best studied is aurora B, a “chromosome passenger protein” that is located on the chromosomes during prophase and becomes concentrated at inner centromeres by metaphase before relocalizing to the spindle midzone at anaphase (Carmena, et al., Nat. Rev. Mol. Cell. Biol. 4:842-854 (2003)). Consistent with this, aurora B has both chromatin and spindle-associated substrates and influences mitosis at a number of steps. Aurora B homologues play an important role in ensuring chromosome bi-orientation at metaphase by correcting mono-orientated attachments to the spindle, and are involved in normal chromatid separation and cytokinesis (Shannon, et al., Curr. Biol. 12:R458-460 (2002); Andrews, et al., Curr. Opin. Cell. Biol. 15:672-683 (2003)). They are also required for phosphorylation of the centromeric histone variant CENP-A at serine-7 and of histone H3 at serine-10 in many organisms (Hsu, et al., Cell 102:279-291 (2000); Adams, et al., J Cell Bio. 153:865-880 (2001); Giet, et al., J. Cell Biol. 152:669-682 (2001); Petersen, et al., J. Cell Sci. 114:4371-4384 (2001); Zeitlin, et al., J. Cell Biol. 155:1147-1157 (2001); Crosio, et al., Mol. Cell. Biol. 22:874-875 (2002); Ditchfield, et al., J. Cell Biol. 161:267-280 (2003); Hauf, et al., J. Cell Biol. 161:281-294 (2003)).
It has not been possible, however, to unambiguously assign the role of mitotic histone H3 serine-10 phosphorylation solely to aurora B (Nigg 2001; Prigent and Dimitrov 2003). Indeed, in Aspergillus, mitotic histone H3 serine-10 phosphorylation is dependent on the kinase NIMA (De Souza, et al., Cell 102:293-302 (2000)). In addition, kinases that bring about the phosphorylation of other histone residues during mitosis must exist. The nature of these key enzymes remains unclear, although there is some evidence that aurora B and the Dlk/ZIP kinase are responsible for mitotic phosphorylation of H3 serine-28 and threonine-11 respectively (Goto, et al., Genes Cells 7:11-17 (2002); Preuss, et al., Nucleic Acids Res. 31:878-885 (2003)).
Haspin/Gsg2 (Haploid Germ Cell-Specific Nuclear Protein Kinase/Germ Cell Specific Gene-2) was first identified as a testis-specific gene in mice (Tanaka, et al., FEBS Letts. 355:4-10 (1994); Tanaka, et al., J. Biol. Chem. 274:17049-17057 (1999)). More recent work has suggested that lower levels of haspin mRNA are also present in other organs and in all proliferating cell lines tested, suggesting that expression of haspin is not truly haploid germ cell-specific (Higgins, Gene 267:55-69 (2001). Genes encoding haspin homologs are present in all major eukaryotic phyla, including yeasts, microsporidia, plants, nematodes, flies, fish, amphibians and mammals (Higgins, Cell Mol. Life. Sci. 60:446-462 (2003)). These haspin genes encode proteins that contain a distinctive C-terminal putative kinase domain and together constitute a novel eukaryotic protein kinase family (Higgins, Prot. Sci. 10:1677-1684 (2001). The N-terminal portion of the haspin proteins is less conserved between species and has no clear homology to known domains (Tanaka, et al., J. Biol. Chem. 274:17049-17057 (1999); Yoshimura, et al., Gene 267:49-54 (2001); Higgins, Cell Mol. Life. Sci. 60:446-462 (2003)).